αβ-TCR+CD4¯CD8¯NK1 .1¯ Double-Negative (DN) regulatory t cells: gene expression profiling and functional analysis

Abstract

The role of alphabetaTCR+CD4-CD8 -NK1.1- double negative (DN) regulatory T (Treg) cells from both mice and humans has been demonstrated to be involved in settings of autoimmunity, transplantation and cancer. The focus of this thesis was to develop a gene expression profile to characterize the molecular mechanisms underlying the observed function of DN Tregs. Analysis of the gene expression profile identified several possible mechanisms which may explain DN Treg antigen detection, migration, IFN response, survival, cytotoxicity, apoptosis and immune modulation. CXCR5 was one of several chemokine receptors found to be up-regulated in DN Tregs. Characterization of CXCR 5 expression and function on DN Treg cells identified an important function in allograft homing. Using a murine cardiac allograft transplant model, we found that early production of CXCL 13 following transplantation preferentially directed DN Treg cell homing allografts. We further demonstrate the role of HSP70 in DN Treg cytoprotection following antibody mediated TCR cross-linking as a mimic for activation induced cell death (AICD). Cytosolic HSP70 protein expression was up-regulated immediately following cross-linking in both DN Tregs and CD8 T cells. However, in DN Tregs this up-regulation was maintained while CD8 T cells rapidly mobilized HSP70 to the cell membrane. We found membrane HSP70 expression was correlated with apoptosis and describe a previously unknown involvement of HSP70 in Treg resistance to AICD. The data presented within this thesis describe important mechanisms responsible for DN Treg tissue homing and survival. Better understanding of molecular mechanisms utilized by DN Tregs for regulatory function may allow us to manipulate them for cell based immunotherapies.